iNOS gene expression is regulated by various pathways including NF-κB, IFN-γ (signaling through the JAK-STAT cascade) and HIF-1, which induce expression, and TGF-β and interleukins -4, -10 and -13, which inhibit expression. Post-translational regulation occurs by prevention of iNOS homodimerization by NAP110.

iNOS has roles in the innate immune system and displays antibacterial, antiviral and antifungal activity. The human gene encoding iNOS is localized to chromosome 17q11.2-q12.